不同血清培养条件对人脐带间充质干细胞增殖及分化的影响

Effects of serum culture conditions on proliferation and differentiation of human umbilical cord mesenchymal stem cells in vitro

目的观察不同血清培养条件对人脐带间充质干细胞（hUC—MSCs）增殖活力和诱导分化为肝细胞的影响。方法用组织块贴壁法分离hUC—MSCs，分别以下列血清培养条件进行培养：分组A：2％UhroserG血清替代品，分组B：1％UhroserG血清替代品＋5％脐血血清，分组C：10％脐血血清，观察细胞的形态特征、增殖能力。利用含肝细胞生长因子（20μg/L）和成纤维细胞生长因子（10μg/L）的培养基将hUC—MSCs向肝细胞诱导分化，免疫组织化学方法检测诱导分化后甲胎蛋白（AFP）、细胞角质素18（CK18）、白蛋白（ALB）蛋白的表达。结果不同血清培养条件下hUC—MSCs的形态正常，2％UltroserG血清替代品组与1％UhroserG血清替代品＋5％脐血血清组的增殖能力均高于单纯10％脐血血清组（分别较单纯血清组高35．8％和17．5％，P〈0．05）。诱导分化后AFP、ALB、CK18蛋白在含UltroserG血清替代品组中的表达高于单纯脐血血清组。结论含血清替代品的培养条件有助于hUC．MSCs的增殖及向肝细胞分化。

Objective To study the effects of different serum culture conditions on the prolifera- tion and differentiation into liver cells of human umhilical cord mesenchymal stem cells （ hUC - MSCs） in vitro. Methods hUC - MSCs were isolated and cultured with tissue explants adherent method. The serum conditions for the culture of hUC - MSCs were 2% Uhroser G serum substitute （group A） , 1% Uhroser G serum substitute ＋5% umbilical cord serum （group B）, and 10% umbilical cord serum （group C）, re- spectively. The morphologic features and growth status of hUC - MSCs were observed by an invert micro- scope. The cell proliferation was quantitated by the colorimetric WST - 8 assay. Then hUC - MSCs of pas- sage 3 （abbreviated to P3 ） were induced into liver cells with hepatocyte growth factor （HGF, 20 μg/L） and fibroblast growth factor 4 （FGF4, 10 μg/L）. The immunocytochemistry was used to detect the expres- sion of AFP, ALB and CKI8 proteins in the induced hUC - MSCs. Results The hUC - MSCs grew well and uniformly in culture. The proliferation ability of hUC - MSCs in group A and group B was higher than that in group C （35.8% vs. 17.5% , respectively）. The expression levels of AFP, ALB and CK - 18 pro- teins in group A and group B were also higher than those in group C. Conclusion The culture conditions containing serum substitute facilitates the proliferation and differentiation into the liver cells of hUC - MSCs in vitro.